Gene Center, LMU Munich
Institute of Molecular Animal Breeding and Biotechnology
Genetic modification of islet donor pigs - what is the evidence for improved efficacy?
Eckhard Wolf1, Nikolai Klymiuk1.
1Gene Center, LMU Munich, Munich, Germany
Recent developments in pig genome engineering facilitate precise and efficient modifications of donor pigs for islet xenotransplantation. Major goals of genetic modifications are to overcome immune rejection of xeno-islets, to foster their survival, engraftment and re-vascularization, and to improve their functionality. The individual requirements of genetic optimization of donor pigs depend on the kind of islet product (neonatal vs. adult) and the transplantation site. If naked islets are delivered intraportally, a significant proportion of islet mass is destroyed by the instant blood-mediated inflammatory reaction (IBMIR), which involves activation of complement and coagulation, and infiltration of innate immune cells. Consequently, islets from pigs lacking the major xenoantigen Gal-α1,3-Gal (GTKO) and/or expressing human complement regulatory proteins were tested in non-human primate models. GTKO was shown to be advantageous for survival and engraftment of neonatal islet cell clusters (NICCs) in streptozotocin-diabetic rhesus monkeys with immunosuppression . However, data from an elegant “dual islet transplant model”, where different islet preparations can be compared within the same recipient, suggested that this was not due to an improved control of IBMIR as no significant differences in complement and antibody deposition, and infiltration by neutrophils, macrophages and platelets were found between GTKO and wild-type (WT) NICCs within the first day post-transplant . In combination with transgenic expression of human (h) CD55 and hCD59, however, GTKO NICCs clearly attenuated IBMIR after intraportal transplantation compared to WT NICCs . While GTKO did - presumably due to the loss of Gal-α1,3-Gal epitopes during postnatal islet development - not influence the outcome of intraportally transplanted adult pigs islets , expression of hCD46 supported their long-term survival, although it had no effect on islet loss in the early post-transplant period . A more recent study evaluated the effects of additional transgenes [hCD39, human tissue factor pathway inhibitor (hTFPI) and porcine (p) CTLA4-lg] expressed under a beta-cell specific promoter on the GTKO/hCD46 background. Additional expression of hCD39 (n=1), hTFPI + pCTLA-4Ig (n=2), or hTFPI + hCD39 + pCTLA-4Ig (n=2) did not show consistent improvement of survival and function of adult islets after intraportal transplantation into streptozotocin-diabetic immunosuppressed cynomolgus monkeys as compared to historical experiments with islets from WT pigs or pigs with only one genetic modification . Differences in the conditions of islet isolation and potential epigenetic effects on transgene expression in cloned multi-modified pigs were discussed as potential limitations of this study. In summary, only few systematic efficacy studies of genetically modified porcine islets have been performed, and comparison of their results is complicated by different experimental conditions and readouts. In addition, high costs and ethical constraints of non-human primate studies limit the number of recipients. New experimental systems, such as the “dual islet transplant model”  have the potential to better clarify whether a specific genetic modification, immunosuppressive treatment, or a combination of both will result in an improved outcome. While so far mainly islets with genetic modifications targeting complement mediated processes such as IBMIR have been tested in non-human primates, transgenic strategies to overcome T-cell mediated islet xenograft destruction, such as beta-cell specific expression of LEA29Y , still require this validation step.
Grant support: German Research Council (Transregio CRC 127).
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 Klymiuk et al., Diabetes 61, 1527-32 (2012)
11:00 - 13:30
|Prospects for Clinical Islet Xenotransplantation (CIX)||Genetic modification - what is the evidence for improved efficacy?||Room 109|